ES2943382T3 - Flow cell for the analysis of particles in a liquid to be examined, procedure and use - Google Patents

Abstract

The invention relates to a device (1, 2) for examining particles (RBC, WBC, PLT) in a liquid to be examined, comprising a flow passage (F) through which the liquid (BL) to be examined moves. . The flow passage (F) has at least one inlet (E3, E4) through which at least one sheath fluid (PF1, PF2) flows towards the flow passage (F) so that at least one fluid is formed envelope (PF1, PF2). at least one sheath flow (MS1, MS2) in the flow passage (F). The device (1, 2) further comprises a wave generating device (WE2) for piezoacoustically generating sound waves (AW) that propagate through the flow passage (F) transversely to the flow direction of the liquid (BL) to examine and form a wave. nodes (KN) in a monitoring plane (BA) so that the particles (WBC, RBC, PLT) to be examined from the liquid (BL) to be examined move to the monitoring plane (BA) and accumulate based on the pressure effect of sound waves (AW) in the transverse direction. (Automatic translation with Google Translate, without legal value)

Description

DESCRIPTION

Flow cell for the analysis of particles in a liquid to be examined, procedure and use

The invention relates to a device for the examination of particles in a liquid to be examined. Furthermore, the invention relates to a device for the microscopic observation of particles in a liquid to be examined. Furthermore, the invention relates to a method for the examination of particles in a liquid to be examined. Furthermore, the invention relates to the use of a flow cell and a wave generating device for the piezoacoustic generation of sound waves.

Blood consists of various components that are present in the blood in different concentrations. Red blood cells, also called erythrocytes, represent 98% of the blood, platelets, also called thrombocytes, on the contrary, comprise only 1.9% of the blood, and white blood cells, also called leukocytes, they even have only a proportion of 0.1% in the blood. When it is desired to examine only the blood components with low ratio, that is, platelets or white blood cells, there are two possible ways of operating. Either red blood cells present at a frequency greater than up to 1000-fold are detected together with the cell type of interest, i.e. thrombocytes or leukocytes, or red blood cells are removed prior to analysis , in order to concentrate the components of interest in the test fluid accordingly.

In the first type of approach, the total analysis time is considerably prolonged, that is to say by a factor corresponding to the ratio between the proportion of red blood cells and the proportion of the type of cell to be analysed. In addition, the amount of data to be processed is extremely high in this regard. In the second type of approach, an operation is necessary before the actual analysis, in which the blood components that are not to be analyzed are removed from the analysis liquid. Only with the second procedure is an acceptable reduction in analysis time achieved. Conventional analysis procedures rely on electrical methods, such as Coulter's principle. The principle of the Coulter counter is based on the measurement of the change in average electrical conductivity between two electrodes. These are immersed in an electrically conductive liquid, which contains particles with a different conductivity than the liquid. The measurement device used during the procedure comprises two measurement chambers. In each of the two chambers, which are separated from each other by a narrow opening, a measurement electrode is arranged. In this regard, the size of the opening is determined by the size of the particles to be examined.

Other conventional analysis procedures are based on optical methods. In this context, flow cytometry is mentioned. In this regard, the cells to be examined in a liquid are analyzed, which individually pass with high speed in an electric field or a beam of light. Depending on the shape, structure or coloration of the cells, different effects result, on the basis of which the properties of the respective cells are determined. This principle is used, for example, in hematology analyzers or fluorescence flow cytometers. In fluorescence flow cytometry, cells doped with a fluorescent marker are sorted into different reagent vessels, depending on the staining.

US 2014/0147860 A1 describes acoustic flow cytometry on biological particles. In particular, document US 2014/0147860 discloses a device for the examination of particles in a liquid to be examined, which exhibits a process step for the separation and location of the particles to be examined, comprising:

- a flow passage, through which the liquid to be examined moves with a first flow rate, with at least one inlet, through which the at least one coating liquid flows with at least at least a second flow rate in the first flow path, such that the at least one coating liquid forms the at least one coating flow in the flow path and the liquid to be examined flows longitudinally with respect to the por at least one coating flow through the flow path,

- a wave generation device for the piezoacoustic generation of sound waves, which propagate through the flow passage transversely to the flow direction of the liquid to be examined and form wave nodes in an observation plane of so that, due to the pressure effect of the sound waves in the transverse direction, the particles of the liquid to be examined to be examined move transversely to the observation plane and are concentrated there, so that the observation plane flows through the at least one coating stream and the particles to be examined are concentrated in the at least one coating liquid,

wherein the device further comprises an actuating element for generating the individual flows of the at least one coating liquid and the liquid to be examined.

In addition, microscopy is also used for the analysis of blood components. In this connection, for example, cells are stained on a slide and observed under a microscope. A high efficiency of the cells to be examined can be achieved when staining of the cells with fluorescent substances and observation by microscopy are combined with flow cytometry.

For the separation of particles to be examined in liquids, processes with highly viscous coating liquids (so-called "sheath floW") and piezoacoustic separation of cells are also used. Such processes are described, for example, in US 2004/0070757 A1, US 2010/0009333 A1, Laurel et al., Chem. Soc. Rev., 2007, 36, 492-506 and A. Nilsson et al., Micro Total Analysis Systems 2002, Kluwer Academic Publishers, Nara, Japan 2002.

However, the mentioned procedures have not been used up to now to concentrate weakly concentrated components of liquids to be examined.

Therefore, it is an object of the present invention to indicate an improved device and a corresponding method for the examination of particles in a liquid to be examined, which is suitable for the analysis of weakly concentrated particles in liquids to be examined.

This object is achieved by means of a device for the examination of particles in a liquid to be examined according to claim 1, a device for the microscopic observation of particles in a liquid to be examined according to claim 11 , a method for the examination of particles in a liquid to be examined according to claim 13 and a use of a flow cell and a wave generating device for piezoacoustic generation of sound waves according to claim 14.

The device according to the invention for the examination of particles in a liquid to be examined exhibits a process step for the separation and location of the particles to be examined. The process step comprises a flow path, through which the liquid to be examined moves with a first flow rate. Such a flow passage is preferably configured so that liquids flowing through the flow passage do so in a laminar fashion. For this, suitable diameters of the flow passage and flow rates or flow velocities of the liquids flowing through the first flow passage are chosen. The flow path can be realized for example by using a conductive element or a tubular constituent element, which is preferably optically transparent. Flow rate should be defined as a volume of liquid flowing through a cross section, per unit of time.

The flow passage comprises at least one inlet, through which in each case at least one coating liquid flows in the flow passage with at least one second flow rate, so that the at least one coating liquid forms the at least one coating flow in the flow passage and the liquid to be examined flows through the flow passage longitudinally of the at least one coating flow. A coating flow should in this context therefore be understood to mean a liquid flow, which surrounds a further liquid flow in at least one direction or flows along one side of the other liquid flow in its flow direction, where the coating flow and the other liquid flow flow in the same direction. However, the casing flow may exhibit a different velocity and flow rate than the other liquid flow surrounding or flowing alongside it. As investment flow liquid, also called investment liquid, so-called buffer solutions are usually used, in which particles to be examined remain durably or are protected from changes by the support liquid.

The device according to the invention for the examination of particles in a liquid to be examined further comprises a wave generating device for the piezoacoustic generation of sound waves. Piezoacoustic generation of sound waves is understood to mean the periodic electrical excitation of piezoceramic oscillating elements. In this connection, the piezoceramic oscillating elements are excited with the help of an electric field into a mechanical oscillation, which in turn causes sound waves, which propagate in the liquid flowing through the flow passage. By arranging reflection elements opposite the wave generating device, the formation of standing waves is achieved. In the simplest case, for this, for example, the side wall of a pipe is sufficient, which surrounds and conducts the liquid flowing through the flow passage.

The sound waves generated with the aid of the piezoceramic oscillating elements propagate through the flow passage transversely to the flow direction of the liquid to be examined and form wave nodes in an observation plane in such a way that, due to the effect of pressure of the sound waves in the transverse direction, move the particles to be examined from the liquid to be examined into the observation plane and are concentrated there. An enrichment of the particles to be examined in the observation plane is achieved because during the displacement of the particles to be examined their specific kinematic behavior under the influence of sound waves is used. For example, compared to the rest of the particles, due to sound waves, particles with relatively large dimensions move faster in the transverse direction, compared to small particles. This difference in speed can be used, for example, so that more large particles are clearly concentrated in the observation plane than unwanted smaller particles.

The particles to be examined are transported in the direction of propagation of the sound waves to the wave nodes, where they are no longer exposed to any forces in the transverse direction and therefore only move in the direction of the flow. An observation plane or an observation zone comprises a narrowly limited section in the transverse direction to the flow direction in the flow passage, in which with a optical observation used for the observation of the particles to be examined can be sharply observed objects, in particular the particles to be examined. The extent of the observation zone in the transverse direction therefore depends on the depth of focus of the used optical observation device. In addition, the position of the observation zone in the transverse direction depends on the adjustment and arrangement of the optical observation device.

In the control of inlets or devices for the movement of liquids, the value of the at least one second flow rate is chosen such that the at least one casing flow exhibits a predetermined cross section, so that the The observation plane or the observation zone runs through the at least one coating flow and the particles to be examined are concentrated in the at least one coating flow or in the at least one coating liquid. The choice or control of the flow rates occurs by means of a control unit which mutually predicts and synchronizes values for the individual flow rates, and controls the drive element for the generation of the individual flows of the coating liquids and of the liquid which is to be examined, corresponding to the determined flow rates.

Thus, the central flow is decentered so that during the separation the particles to be examined can be moved from the central flow to a preferably highly transparent coating flow and be there simultaneously in the observation plane or in the observation area. The particles to be examined, separated in this way, can then be easily observed in the preferably highly transparent coating flow downstream, with a corresponding observation device, without the remaining particles in the central flow being able to disturb or even prevent the view of the particles to be examined. With the aid of the device according to the invention for the examination of particles in a liquid to be examined, an enrichment of particles to be examined is thus achieved in an observation plane. In addition, with the help of acoustic waves, an alignment of the particles to be examined can also be achieved corresponding to the orientation of the observation plane, so that they can be well observed. For the alignment of the particles, a propagation direction of the acoustic waves is chosen, perpendicular to the observation plane. In this way, the addition of polymeric substances for the alignment of the particles to be examined, as is conventionally applied, can be dispensed with, or at least the duration during which the particles to be examined are exposed to the polymers can be reduced, in order to their alignment. Since the polymeric substances mentioned can undesirably change their external shape due to a very long-lasting effect on the particles to be examined, it is particularly advantageous if their use can be limited or even completely dispensed with. In this way, the quality of the particles to be observed can thus be preserved and, nevertheless, an optimal alignment of these particles can be achieved, so that they are aligned corresponding to an orientation of an observation plane.

The fact that the particles to be examined can be lifted from the particle flow to another plane, and thereby removed from the liquid flow to be examined, so that the components in the main flow are not have no interfering influence on the observation of the particles to be examined, it can be very advantageous for that purpose, for example during blood examination, that contrary to conventional procedures for examination, undiluted blood which exhibits a higher viscosity than diluted blood. Thus, however, it is particularly suitable for laminar flows, as is the case with the device according to the invention described. With laminar flows, mixing of the coating liquids with the central flow does not occur, so that a separation of particles from the central flow can be carried out particularly effectively.

The device according to the invention for the microscopic observation of particles in a liquid to be examined comprises a device according to the invention for the examination of particles in a liquid to be examined. Furthermore, the device according to the invention for the microscopic observation of particles in a liquid to be examined has a microscope, which is arranged in a longitudinal position of the first flow passage for the observation of particles of the liquid to be examined in the observation plane. Such a microscope can in particular be an optical microscope, but it can also be an electron microscope or a microscope of another type suitable for the observation of liquids. With the microscope, for example, individual particles to be examined can be identified and counted. In addition, with the aid of the microscope, the particles to be examined can be differentiated or classified. For example, leukocytes, thrombocytes and erythrocytes can be differentiated. In addition, a classification of the types of particles observed can also be made, in subspecies. For example, leukocytes can be classified according to five different subtypes.

Details of the particles to be examined can also be observed. Due to the separation of the particles to be examined and the displacement of these particles to the observation plane or the observation zone of the microscope, the observation can be limited to the particles to be examined, without interfering particles impeding the view . Since interfering particles do not occur or occur only to a limited extent during an observation of a liquid containing the particles to be observed in the observation zone, they do not have to be further examined and possibly discarded, so that It clearly facilitates an observation and analysis of the liquid containing the particles to be observed and, due to the low number of the particles to be verified, it is remarkably accelerated.

The particles to be verified are moved in a focused manner by means of the acoustic force or the force of the sound waves, to the examination plane or observation plane of a microscope, in order to enrich these there or impoverish the smallest particles, as for example erythrocytes or thrombocytes with respect to large particles, particularly leukocytes. In this way an enrichment of, for example, 200-2000 fold of the leukocytes can be achieved.

During the method according to the invention for the examination of particles in a liquid to be examined, a liquid to be examined is moved with a first flow rate, through a flow passage. Furthermore, at least one coating liquid with at least a second flow rate is introduced into the flow passage through a first inlet, so that the at least one coating liquid forms a first coating flow in the flow passage and the liquid to be examined flows longitudinally of the at least one coating flow, through the flow passage. In addition, sound waves are generated piezoacoustically. The sound waves propagate through the flow passage, transversely or through the flow direction of the liquid to be examined and form a wave node in an observation plane so that due to the pressure effect of the Sound waves in the transverse direction move the particles to be examined from the liquid to be examined into the observation plane and are concentrated there. In this regard, the value of the at least one second flow rate is chosen such that the at least one casing flow exhibits a predetermined cross section such that the observation plane runs through the at least one flow rate. coating flow and the particles to be examined are concentrated in the at least one coating liquid. Particles displaced into the observation plane or into the observation zone to be examined can then easily be observed with an optical observation unit, such as a microscope.

During the method according to the invention there occurs a use of a flow cell and of a wave generating device, for the piezoacoustic generation of sound waves, in particular of a device according to the invention for the examination of particles in a liquid to be examined, for the separation of leukocytes from a blood plasma. As already mentioned, during the conventional observation of leukocytes, the problem of the presence of these only in low concentration in the blood occurs. An observation or examination of the leukocytes without their separation therefore leads in this respect to the fact that the red blood cells are also considered and have to be discarded. Advantageously, during the use of the device according to the invention, use is made of the fact that the white blood cells move faster under the influence of sound waves, compared to the other components of the blood. By using this fact, white blood cells transversely to the flow direction can now be replaced and separated with the aid of sound waves in a lengthwise flowing blood plasma. The observation of the low, compared to the large number of red blood cells, of separated white blood cells is subsequently carried out with very little effort, so that the observation time is advantageously significantly reduced.

The essential components of the device according to the invention for the examination of particles in a liquid to be examined can be configured predominantly in the form of software components. This concerns in particular parts of the wave generating device, such as a control unit for controlling the generation of sound waves. In addition, this also refers to control units that are used for the control of the flow rates of the liquids that are to flow through the inlets of the flow passage.

Basically, these components can sometimes also be realized, in particular when it comes to particularly fast computations, in the form of software-supported hardware, for example FPGAs or the like. In addition, the necessary interfaces, for example when it is only a question of transferring data from other software components, can be configured as software interfaces. However, they can also be configured as constituent hardware interfaces, which are controlled by suitable software.

An embodiment implemented extensively by software has the advantage that hitherto used control units can also be easily updated by means of a software update to work in the manner according to the invention. In this respect, the object is also achieved by a corresponding computer program product with a computer program, which can be loaded directly into a storage device of a device control device according to the invention, for examination. of particles in a liquid to be examined, with program sections, for carrying out all the steps of the method according to the invention, when the program is carried out in the control device. In addition to the computer program, such a computer program product may optionally comprise additional components, such as documentation and/or additional components, as well as hardware components, such as hardware keys (keys, etc.), for the use of the software.

For transport to the control device and/or for storage of or in the control device, a computer-readable medium, for example a memory stick, hard disk or other fixed or transportable data carrier, on which they store the program sections of the computer program that can be read and performed by a computer unit of the control device. For this purpose, the computing unit can, for example, have one or more microprocessors working together, or the like.

The dependent claims as well as the following description each contain advantageous configurations and developments of the invention. In this regard, in particular the claims of a category of claims may also be developed in a manner analogous to the dependent claims of another category of claims. Furthermore, within the scope of the invention, the different features of different embodiments and claims can also be combined to give new embodiments.

In a configuration of the device for the examination of particles in a liquid to be examined, the flow passage exhibits a first inlet through which a first coating liquid with a second flow rate flows into the flow passage. Furthermore, the flow passage comprises a second inlet opposite the first inlet, through which a second coating liquid with a third flow rate flows in the flow passage. The first inlet and the second inlet are arranged oppositely in such a way that the first coating liquid forms a first coating stream in the flow passage and the second coating liquid forms a second coating stream in the flow passage, so that the liquid to be examined flows between the first coating flow and the second coating flow through the flow path. The values of the second flow rate and the third flow rate are chosen such that the casing flows exhibit a predetermined cross section, such that the observation plane runs through one of the two casing flows. With the aid of the two coating flows, a position of the central flow of the liquid to be examined can be set, either in the cross section of the flow passage or in the transverse direction to the flow direction. In particular, it can be achieved by this that the central flow no longer flows into the observation plane, so that the particles to be examined can be lifted from the central flow into the observation plane. The use of two coating streams allows a free location of the central stream, as well as greater flexibility in the choice of the viewing plane to which the particles to be examined are transferred, where they can be viewed with the aid of a viewing apparatus. such as a microscope.

In a configuration of the device according to the invention for the examination of particles in a liquid to be examined, the flow passage is dimensioned so that the liquids flowing through it flow laminarly. This can be achieved for example when the flow path comprises a microfluidic channel. Whether a laminar flow prevails in the flow passage depends on the diameter of the flow passage and on the speed of the liquids flowing through it and their viscosity. For a given flow rate and viscosity, the diameter of the flow passage is therefore chosen accordingly, so that the liquids passing through the flow passage can pass through it laminarly. The presence of a laminar flow has the advantage that turbulence does not occur in the flow passage, which could lead to an undesired mixture of central flow and coating flow.

In a preferred configuration of the device according to the invention for the examination of particles in a liquid to be examined, the liquid to be examined comprises blood plasma. As already mentioned, the device according to the invention for the examination of particles in a liquid to be examined is particularly well suited for the separation and subsequent observation of leukocytes, which occur in relatively low concentration in blood plasma.

Preferably, with the device according to the invention for the examination of particles in a liquid to be examined, the position of the nodes of the acoustic waves is fixed in the transverse direction, in order to adjust the frequency of the sound waves. The position of the nodes in the transverse direction with respect to the flow direction depends on the wavelength of the standing waves and thus on the frequency used for the excitation of the waves. In this way, the position of the wave nodes is adjusted to a desired position of an observation plane or a central area of an observation area, in the direction transverse to the flow direction.

In a particularly preferred variant of the device according to the invention for the examination of particles in a liquid to be examined, the particles to be examined display leukocytes or erythrocytes or thrombocytes. The described enrichment procedure is also applied for the enrichment of erythrocytes. Red blood cells are 5 to 6 µm in size and are twice the size of thrombocytes, which is approximately 1.5 to 3.5 µm. In this way, with the aid of sound waves, the erythrocytes are also separated from the thrombocytes. In addition, an additional process step could also be connected in series to the described process step, which further separates the leukocytes from the erythrocytes and functions analogously to the described process step.

The thrombocytes can be separated in the manner described above thereby as smaller particles, as a result of the separation of all blood components, i.e., the leukocytes and the erythrocytes, and the diversion of the central flow with the thrombocytes to the plane of observation.

In particular, preferably the device according to the invention for the examination of particles in a liquid to be examined has a separation passage connected in series with the flow passage. In this respect, the separation step exhibits an additional flow step with a central flow containing a liquid, initially free of the particles to be examined, and at least one overlay flow, referred to below as the overlay flow of the separation step. separation, which flows longitudinally of the core flow through the flow passage of the separation passage, and exhibits the liquid to be examined. Preferably, the at least one overlay flow from the separation passage surrounds the central flow. In this context, a central flow should be considered a liquid flow that flows alongside the at least one other liquid flow, in this case a coating flow. How I know As described, the central flow is preferably surrounded by the at least one coating flow so that it then flows centrally through the flow passage.

In this variant, the separation passage also has an additional wave generation device for the piezoacoustic generation of sound waves, which propagate through the flow passage transversely to the flow direction of the liquid to be examined. of the separation passage, and form a wave node in a plane through the central flow so that, due to the pressure effect of the sound waves in the transverse direction, it displaces particles, in particular particles with a larger diameter compared to other types of particles, in the liquid to be examined, to the central flow, and there they are concentrated. In this particularly preferred variant, an additional separation of the particles to be examined, from the liquid to be examined, therefore occurs before the separation of the particles to be examined and their displacement to the observation plane, so that the effectiveness and reliability of the separation of the particles to be examined is raised.

In a particularly advantageous variant of the device according to the invention to be applied for the examination of particles in a liquid to be examined, the liquid initially free of the particles to be examined is transparent. The high transparency of the liquid free of the particles to be examined allows a simplified observation of particles to be examined transferred to this liquid.

In a particularly advantageous configuration of the device according to the invention for the examination of particles in a liquid to be examined, the wave generation device of the separation stage and of the process stage is configured such that the direction of propagation of the sound waves in the separation passage and the direction of propagation of the sound waves in the flow passage of the process passage run orthogonal to each other, and orthogonal to the flow direction of the liquid to be examined .

Alternatively, the separation step and the process step are configured such that the separation in the separation step and the subsequent displacement of the central flow in the process step run in the same direction.

For this, the dimensions of a channel defining the flow passage or flow path can be changed, or the wave generating device can be positioned laterally, and not vertically.

The alignment of the propagation direction of the acoustic waves is determined by the geometric shape of the conduction elements that surround the flow path, and by the alignment and position of an observation unit that will be assigned to it, such as an observation microscope.

In a special variant of the device according to the invention for the examination of particles in a liquid to be examined, the flow passage of the process passage is enlarged before the longitudinal position of the wave generating device of the process passage , transversely to the flow direction, so that the flow velocity of the liquid to be examined and of the at least one coating liquid are reduced. As a consequence of the lower flow velocity, the particles to be examined are more easily observed and analyzed, since they remain longer in which the particles are kept in the observation zone of a used observation apparatus.

Preferably, the device according to the invention for the examination of particles in a liquid to be examined has a buffer addition unit. The buffer addition unit is arranged to add a buffer to the hypotonic solution of the core flow or the fluid to be examined. With the help of the additional buffer, particles with a high intracellular osmotic pressure are dissolved, while particles with a low intracellular osmotic pressure remain.

During the separation of white blood cells from a blood plasma, red blood cells dissolve with the aid of the additional buffer solution due to their high intracellular osmotic pressure, while the white blood cells remain. This measure thereby makes possible a greatly improved separation of the white cells to be examined from the blood, from the blood plasma.

Particularly preferably, the device according to the invention for the examination of particles in a liquid to be examined exhibits a unit which is preferably connected to the second input and is adapted to add markers for the target particle to at least a coating liquid. Markers can have specific chemical properties such that they bind only certain components to be tested in a liquid to be tested. For example, they can display a special color and thus easily mark the components of interest in the liquid to be examined. The use of markers therefore facilitates the traceability of the particles to be observed in a liquid to be examined. The markers can also have physical properties which facilitate or also make possible a separation of the particles attached to them. For example, basically difficult-to-separate particles could be provided with suitable markers, so that the separation procedures described above are more easily and effectively applicable. Occasionally a situation may occur in which different particles present in a liquid are almost the same size and are therefore only hardly separated from each other, with waves of sound. If the particles of interest are now provided with markers of correspondingly large dimensions, a separation of the particles of interest can nevertheless take place with the aid of sound waves.

Furthermore, the device for microscopic observation of particles in a liquid to be examined may exhibit a collection unit, which is arranged in series connection to the process step, downstream of the longitudinal position of the microscope and is adapted to collect the particles. particles under study. The collected particles can then be further examined with another examination method, which requires an enrichment of the examined particles, preferably molecular biology methods. In this way, the targeted separation of particles to be examined and enrichment can be used several times, successively and for different examinations, with the aid of the device according to the invention.

The device according to the invention for the microscopic observation of particles in a liquid to be examined can furthermore have a particle transfer unit, which is arranged downstream, i.e. connected after the process step, behind the longitudinal position of the microscope, and configured for transfer of the particles, particularly the particles under examination, from the flow path to a slide, for further examination of the particles. Such a transfer unit can for example be part of the described collection unit and exhibit an opening through which drops of concentrated liquid move with the particles to be examined, so that they then fall onto a slide. The slide can be used as a means of transporting the particles under examination to another position, in which other examinations are performed.

The invention is illustrated once again in more detail below with reference to the accompanying figures, by virtue of the exemplary embodiments.

FIG 1 shows a schematic representation of a device for the examination of white blood cells according to an exemplary embodiment of the invention,

FIG 2 shows a schematic representation of a white blood cell separation process in one separation step,

FIG 3 shows a schematic representation of a displacement of white blood cells to an observation plane,

FIG 4 shows an overview of the separation and displacement procedures of a device for the examination of white blood cells, during the application of a hypotonic solution according to an exemplary embodiment of the invention,

FIG 5 shows a schematic representation of a device for the examination of white blood cells according to an exemplary embodiment of the invention, with a slide connected in series,

FIG 6 shows a flowchart, with which the course of a procedure for the examination of white blood cells is illustrated, according to an exemplary embodiment of the invention.

A first exemplary embodiment of a device 1 for the examination of WBC white blood cells is described with reference to FIGS. 1 to FIG. 3.

In FIG. 1, a top view of a microfluidic channel system 1 is shown, in which a device for examining particles in a liquid to be examined, according to an exemplary embodiment of the invention. The microfluidic channel system comprises a separation step S1, a process step S2 connected in series with the separation step and a collection unit S3 following the process step. The top view occurs on the x/y plane. In this regard, the flow direction of the liquid to be examined is the x-direction. Additionally, in FIG 1 the part presented on the right side is represented, that is, the process step S2, which comprises a flow step F, and the collection unit S3 of the microfluidic channel system 1, bottom right. in the side view image, that is, in the x/z plane.

Microfluidic channel system 1 exhibits a separation step S1. The separation step S1 comprises a first inlet E1, in which the blood plasma BL flows with a first flow rate v ¹ in the microfluidic channel system 1. Additionally, the microfluidic channel system 1 comprises a second inlet E2, through which a blood plasma-free PF fluid, in this case a buffer PF fluid, flows with a second flow rate v ² . The two liquids BL, PF are combined in a flow passage F1 or a first flow passage section F1, and even so that the buffer liquid PF forms a central flow ZS and the blood plasma BL forms a lining flow of the passage Separation MS around the buffer liquid PF, which flows along both sides outside the central flow ZS. Due to the small dimensions of the microfluidic channel 1 (the width is between 100 and 1500 um and the height is about 30 to 500 um) the central ZS flow and the liner flow of the separation MS pass behave, for typical velocities flow, in each case laminar so that they flow past one another, without mixing. The blood plasma BL to be examined flows in the central flow ZS on both sides along and after passing the first flow passage section F1, it finally returns to a zone SB drain, which in FIG. 1 is drawn on the side of the first input E1 opposite the second input E2, that is, completely to the left. In the exemplary embodiment shown in FIG. 1, the blood plasma BL comprises red blood RBC cells, white blood WBC cells and PLT platelets.

Also arranged in the first flow passage section F1 is a first wave generating device WE1 for the piezoacoustic generation of sound waves AW (see FIG. 2). The first wave generation device WE1 is symbolized in FIG 1 with a star. With the aid of the first wave generating device WE1, sound waves AW are generated which propagate perpendicular to the flow direction (see FIG. 2), which form wave nodes in a central region of the first flow passage section F1 , in which the central ZS flow runs. The WBC white blood cells, which are suspended in the lining flow of the separation passage MS, are deflected by the sound waves AW in the direction of the central flow ZS, due to their size, with greater speed than other components of the blood. BL blood, such as RBC red blood cells or PLT platelets. Thus, in the first flow passage section F1, a first enrichment of WBC white blood cells occurs in the central flow ZS.

In the other process course PR there now occurs a transition from the central flow ZS to a process step S2 of the microfluidic channel system 1. The process step S2 comprises a flow step F with a second F2, a third F3 and a fourth flow section F4. In the second flow section of the flow path F, a damping liquid PF1 with a third flow rate v3 flows from a third inlet E3 and from a fourth inlet E4 in the opposite z-direction to the third inlet E3, a liquid PF2 flows. damper with a fourth flow rate v4 to the second flow passage section of the flow passage F. The two buffer liquids PF1, PF2 in each case form a first or a second lining flow MS1, MS2 and limit the central flow ZS with the WBC white blood cells to a region lying between the two lining flows MS1, MS2 , which is not in an observation plane or an observation zone BA of a microscope M subsequently used for observation (see also FIG 2). This flow structure is achieved in a third flow passage section F3. In the third flow passage section F3, the microfluidic channel 1 widens in the y direction, ie transversely to the flow direction, in order to reduce the speed of the first and second coating flows MS1, MS2 and of the central flow ZS. A slowdown of the flow rates is necessary in order to subsequently be able to observe individual white blood WBC cells under an M microscope. As recognized in a magnified representation in FIG 1, the position of the central flow ZS depends on the magnitude of the flow rates v3, v4 of the first and second lining flows. If v4 < v3, then the central flux is shifted in the z-direction upward. If v4 = v3, then the central flow ZS is, viewed in the z-direction, exactly in the center of the cross section of the flow passage F, and if v4 > v3, then the central flow ZS is, viewed in the z-direction, by below the center of the cross section of the flow passage F.

In a fourth flow passage section F4, with the aid of a second wave generating device WE2 (symbolized in FIG. 1 as a star), propagating sound waves AW are again generated perpendicular to the flow direction, without However, in this case they propagate in the z direction and not in the y direction. Sound waves AW, which are preferably standing waves, again exhibit wave nodes KN, which, however, this time are not in the flow zone of the central flow ZS, but are adjacent to the flow zone of one of the two flows MS1, lining MS2 (in FIG 3 the nodes KN are in the first lining flow MS1). The wave nodes are, however, in the viewing plane BA or, viewed in the z-direction, at the level of the viewing area BA of the microscope M. Due to the size differences already illustrated, now again the white WBC cells of the blood are preferably deflected into the observation plane BA, i.e. they arrive at the first coating stream MS1 (see FIG 3), through which the microscope observation plane BA of the microscope M also runs. The first coating stream MS1 is preferably colorless, so that the white blood WBC cells found in the first coating MS1 flow are well recognizable. In an observation section B arranged at the end of the fourth flow passage section F4, observation now occurs perpendicular to the flow direction of the WBC white blood cells through a microscope M located perpendicular to the direction ie, aligned in the z-direction, in the fourth flow passage section F4. For example, with the aid of the microscope M the white blood cells WBC can be counted and/or their structure and construction can be observed. Next, in a third section of the microfluidic channel system 1, also referred to as collection zone S3, a collection of the white blood WBC cells occurs, which can then be used for further examinations.

In FIG 2 the separation process of the WBC white blood cells in the first flow section F1 of the separation step S1, and their enrichment in a central flow ZS comprising a buffer solution, is shown magnified. As already illustrated, the white blood cells WBC, which are in the lining flow of the separation passage MS, are deflected with the aid of sound waves AW in the direction of the central area of the first section F1 of the separation passage. flow where, due to the arrangement of the nodes KN of the sound waves AW, they remain on the central axis of the first flow passage section F1 in the central flow ZS. KN nodes are indicated by intersecting lines. The sound waves AW are reflected at the boundary side of the first flow passage section F1 opposite the sound wave generating device WE1 and form standing waves, which form the outlined wave nodes. Acoustic AW waves make it possible for larger cells, in this case WBC white blood cells (approximately 7-15 um in diameter), to move faster towards the central zone of the first F1 section of passage flow, compared to smaller cells, such as RBC red blood cells (approximately 5 to 6 um in diameter). In the central zone of the first section F1 of flow passage, where the nodes KN of the waves are located. AW acoustically, the white blood cells WBC are moved only by the central laminar flow ZS and thereby move along the central axis of the downstream microfluidic channel 1, ie in the process direction PR. During the repositioning of the WBC white blood cells perpendicular to the flow direction, the WBC white blood cells are transferred to another medium, the ZS central flow medium. The central ZS flow is free of other particles and is therefore highly transparent. The diameter of the coating flow of the separation passage MS and the central ZS flow or its position as well as the enrichment of the WBC white blood cells in the central ZS flow can be influenced with the aid of the flow rates v ¹ or ov ² of the flow MS of coatings or of the flow ZS central. For a ratio of the first flow rate v ¹ to the second flow rate v ² of 10, an enrichment of WBC white blood cells in the central ZS flow is achieved, which on the other hand is associated with a strong reduction in the proportion of red blood cells in the central ZS flow, compared to the blood plasma BL in the lining flow of the separation step MS.

As already mentioned, the central flow ZS flows further downstream to a process step S2 and there flows into a second flow passage section F2 in a third and a fourth inlet E3, E4 along, of which the first and second MS1 flows, coating MS2 flows with third and fourth rates v3, v4 flow to microfluidic channel 1. The first and second two lining streams MS1, MS2 control the location of the central stream ZS in the z-direction. This scenario is magnified in FIG 3. On the one hand, the extension of the central flow ZS, which exhibits a flow rate vs, is limited by the first and second lining flows MS1, MS2 in z-direction, on the other hand, the location of the central flow ZS is fixed with the help of the choice of the third and fourth with flow rates V ³ , V ⁴ of the two casing flows MS1, MS2, so that the central flow ZS is outside of an observation plane or an observation zone BA of a microscope M, which is arranged in an observation position or an observation section B of the microfluidic channel 1. This central flow location ZS occurs in a third flow passage section F3. The flow rate of the central flow ZS, initially denoted by v ² , obviously remains the same, although the cross section or cross-sectional area of the central flow ZS changes after the second flow passage section F2. To indicate this, from the third flow step section F3 the reference sign "vs" is used. The cross section of the central flow ZS depends, from the third flow passage section F3, on the flow rates of the first and second lining flows MS1, MS2.

In order to further concentrate the white blood WBC cells and to keep the red blood RBC cells still present in the central flow ZS outside the observation zone BA, with the aid of the first and second coating flows MS1 and MS2 the Central ZS flow in a non-centred position of microfluidic channel 1.

In the other process course PR, in a fourth flow passage section F4, with the aid of a second wave generation device WE2, acoustic waves AW are generated with a direction of propagation transverse to the flow direction and nodes KN of waves in the plane of observation or in the zone ^b A of limited observation in the z direction. Thus, as a result of the pressure of the sound waves AW, the larger white blood cells WBC are displaced into the first investing flow MS1 and thus into the observation plane BA, while the larger red blood RBC cells small ones remain predominantly in the central ZS flow. Due to the significantly higher strength of the acoustic AW waves, compared to the flow forces of the central ZS stream and of the two lining MS1, MS ² streams, the WBC white blood cells have sufficient time to change to the primer in coating flow MS1, before entering observation section B. There the WBC white blood cells can be easily observed in the transparent PF1 buffer of the first coating flow MS1. Due to its limited depth of focus, the microscope M exhibits a limited observation zone BA in the z-direction. Since the WBC white blood cells are transferred exactly to this observation zone BA, they can be observed with optimum resolution.

In FIG 4 the two-step separation and placement process illustrated in FIGS 1 to FIG 3 is illustrated, again by a sequential representation of the two separation and placement procedures, where for the exemplary embodiment according to FIG 4 additionally adds a hypotonic type damper for a first flow passage section F1 in the central flow ZS. While in the first flow passage section F1 a first enrichment of WBC white blood cells occurs in a central flow ZS, by displacement of the WBC white blood cells in the y direction, and these remain unaffected by the flow as well. hypotonic buffer, the red blood cells, which unintentionally enter the central ZS flow from the lining MS flow of the separation passage, are gradually dissolved by the hypotonic conditions in the central ZS flow, so that the concentration of red blood cells gradually decreases in the central ZS flow. This is clarified by the hatching of an arrow K ^e symbolizing the concentration of red blood cells, where the larger spacing between the hatch lines should signify a lower concentration of red blood cells.

This central flow ZS is then decentered in the z-direction in a second flow passage section F2 (see FIG. 1). After the broadening of the fluid flows ZS, MS1, MS2 in the y-direction, white blood cells are separated again in a fourth flow section F4 (see FIG 4 on the right side), however in this case from the central flow ZS in the za-direction a first lining flow MS1. There, the white blood cells WBC are kept in a zone z corresponding to the observation zone BA and then arrive downstream at the longitudinal position B (see FIG 1), in which a microscope M is arranged. Also during the step of the fourth flow section F4 red blood cells possibly still present in the central flow ZS are dissolved again. As already mentioned, with the help of the arrow KE running in process PR direction, symbolizes the increasingly low downstream concentration of red blood cells in the central flow ZS, so that shortly before microscope position B M in the observation zone BA almost only white blood WBC cells are present. In addition, colored markers, such as SDS, can also be added to the first coating flow MS1 via the fourth entry E4 in the second flow path section F2, with which the morphology of the cells is changed.

Illustrated in FIG 5 is a schematic representation of a microfluidic channel system 2, for the examination of WBC white blood cells according to an exemplary embodiment of the invention with a serially connected OT slide. The construction of the microfluidic channel system 2 shown in FIG 5 largely corresponds to that of the system shown in FIG 1. Additionally, the system 2 shown in FIG 5 however still comprises a slide OT, onto which they fall (force of gravity in x-direction) D ^wbc drops with a defined volume of, for example, 20 to 35 pl, from the collection zone S3 of the microfluidic channel system 2, which contain WBC white blood cells. The OT slide can be moved in the MD direction of movement, in this case in the y direction, so that the individual drops D ^wbc with the white blood cells WBC y arrive successively and in order on the OT slide. The D ^wbc drops on the OT can then be processed using various standard staining technologies for cell identification, such as Hoechst staining. As a consequence of the strong enrichment of WBC white blood cells with the aid of the method according to the invention, it becomes possible, for example, to identify WBC white blood cell pathologies with a much higher statistical significance, compared to the case, for example currently with the aid of a Giemsa staining of white blood WBC cells, in a non-concentrated blood sample.

In FIG. 6 a flowchart 600 is shown, with which the course of a procedure for the examination of white blood cells according to an exemplary embodiment of the invention is illustrated. During step 6.I blood plasma BL is introduced into a separation passage S1 via a first inlet E1 into a peripherally arranged inflow of a first flow passage section F1 of a microfluidic channel system 1. In step 6.II a second inlet E2 of the microfluidic channel system 1 is opened through which a buffer solution PF is introduced into a centrally arranged inflow of the first flow passage section F1. While the blood flowing through the first inlet E1 in the first flow passage section F1 forms an overlay flow of the separation passage MS with a first flow rate vi, the buffer PF flowing centrally through the flow passage Flow step F1 forms a central flow ZS with a second flow rate v ² . In step 6.III, sound waves A ^w are now generated by a sound wave generating device, for example with the aid of piezo elements, which propagate transversely to the flow direction of the first flow passage section F1 of microfluidic channel system 1. As a consequence of the pressure arising from the standing sound waves AW in the transverse direction of the first flow passage section F1, white blood cells WBC are increasingly transferred from the lining flow MS of the separation passage into the flow central ZS, so that they are enriched in the buffer PF solution that forms the central ZS flow. In step 6.IV, after the transition to a process step S2 with a flow step F in a second flow section F2 of the flow step F, a third input E3 and a fourth input E4, are now controlled in step 6.IV, so that highly transparent buffer solutions PF1, PF2 flow into the second flow passage section F2 of the microfluidic channel system 1 and form first and second coating flows MS1, MS2 surrounding the central flow ZS in z-direction. In this regard, the flow rates v3, v4 of the first and second lining flows MS1, MS2 are chosen such that the central flow ZS is decentered in the z-direction, i.e., in the direction of microscope observation, i.e., the central flow ZS is displaced in the z-direction, so that a posterior BA plane of observation no longer flows.

After the flow passage F has been enlarged by a third flow passage section F3, so that the flow velocity of both the central flow ZS and also the first and second coating flows MS1, MS2 decreases in a stage 6 .V in a fourth flow passage section F4 of the flow passage F in a direction transverse to the flow direction, sound waves AW are generated again, which propagate transversely to the flow direction of the passage section F4 of flow step F flow of microfluidic channel system 1. As a consequence of the pressure arising from the standing sound waves AW in the transverse direction of the flow passage section F4, white blood cells WBC are increasingly transferred from the decentered central flow ZS to one of the first and second flows MS1, MS2 coating. The sound waves AW form standing waves, the wave nodes of which are positioned in the z-direction so that, viewed in the z-direction, they are arranged in the height or in an observation plane, which runs through an observation zone BA of a microscope M arranged downstream. In a step 6.VI the white blood WBC cells pass a longitudinal position B of the microfluidic channel system 1, in which a microscope M is arranged. The white blood WBC cells located in the z-direction in the area BA observation points of the microscope M are now observed with the microscope M. After observation, the WBC white blood cells are collected in a stage 6.VII in a collection unit S3 and are subsequently dispensed onto a slide OT in a stage 6.VIII, in the form of drops for other analyses.

To conclude, it is indicated once again that the procedure and device described above are only preferred embodiments of the invention and that the invention can be varied by the expert, without losing the scope of the invention, as long as the specifications are specified. claims. For example, channel arrangement 1 of microfluidics, in the context of the observation and analysis of WBC white blood cells. However, the invention is not limited to the separation and observation of white WBC cells from the blood, but can also be applied to other blood cells or also to fluids other than blood, and to particles occurring in these fluids. For the sake of completeness it is also indicated that the use of the indefinite article "a" or "one" does not exclude that the traits in question may also be present several times. Likewise, the concept of "unit" does not exclude that it consists of several components that, if necessary, can also be spatially distributed.

Claims (14)

1. Device (1, 2) for the examination of particles (WBC, RBC, PLT) in a liquid (BL) to be examined, which exhibits a process step (S2) for the separation and location of the particles that are to be examined (WBC, RBD, PLT), which includes: - a flow path (F), through which the liquid (BL) to be examined is moved with a first flow rate (vs), with at least one inlet (E3, E4), through which at least one coating liquid (PF1, PF2) flows with at least a second flow rate (v3, v4) into the first flow step (F), such that the at least one liquid (PF1, PF2) of coating forms at least one flow (MS1, MS2) of coating in the flow path (F) and the liquid (BL) to be examined flows longitudinally of the at least one flow (MS1, MS2) lining, through flow passage (F), - a wave generation device (WE2) for the piezoacoustic generation of sound waves (AW), which propagate through the flow passage (F) transversely to the flow direction of the liquid (BL) to be examined , and form wave nodes (KN) in an observation plane (BA), so that due to the pressure effect of the sound waves (AW) in the transverse direction, the particles (WBC, PLT, RBC) that go to be examined from the liquid (BL) to be examined are moved transversely to the observation plane (BA) and are concentrated there, wherein the value of the at least one second flow rate (v3, v4) is chosen such that the at least one liner flow (MS1, MS2) exhibits a predetermined cross section such that the plane (BA ) of observation runs through the at least one coating stream (MS1, MS2) and the particles (WBC, PLT, RBC) to be examined are concentrated in the at least one coating liquid (PF1, PF2) , wherein the device is configured by means of an arrangement of reflection elements opposed to the wave generation device (WE2) for the formation of standing waves, wherein the device further comprises actuation elements for the generation of individual flows of at least one coating liquid (PF1, PF2) as well as the liquid (BL) to be examined, wherein the device further comprises a control unit for choosing or controlling flow rates, wherein the control unit is configured to predict and synchronizing the values for the individual flow rates and controlling the drive element to generate the individual flows of the coating liquids and the liquid to be examined corresponding to the determined flow rates. Device (1,2) according to claim 1, wherein the flow passage (F) exhibits a first inlet (E3), through which a first coating liquid (PF1) flows at a second rate. (V ³ ) of flow in the passage (F) of flow, and a second entrance (E4) opposite to the first entrance (E3), through which flows a second liquid (PF2) of coating with a third rate ( V ⁴ ) of flow in the flow path (F), so that the first coating liquid (PF1) forms a first coating stream (MS1) in the flow path (F) and the second liquid (PF2) of coating forms a second coating stream (MS2) in flow passage (F) and the liquid (BL) to be examined flows through flow passage (F) between the first coating stream (MS1) and the second flow (MS2) coating, where the value of the second flow rate (V ³ ) and of the third flow rate (V ⁴ ) is chosen such that the coating flows (MS1, MS2) exhibit a predetermined cross section, such that the plane (BA) of observation runs through one of the two coating streams (MS1, MS2). Device according to claims 1 or 2, wherein the flow passage (F) is sized so that the liquids (BL, PF1, PF2) that flow through it do so in a laminar manner, and/or the flow passage (F) comprises a microfluidic channel (MFK). Device according to one of claims 1 to 3, wherein the liquid (BL) to be examined comprises blood plasma and/or the particles (WBC, RBC, PLT) of the liquid (BL) to be examined ) to be examined exhibit at least one of the following blood components: - leukocytes, - thrombocytes - erythrocytes. Device according to one of claims 1 to 4, having a separation stage (S1) connected upstream to the process stage (S2), wherein the separation stage (S1) comprises: - a flow path (F1) with a central flow (ZS) with a liquid (PF) initially free of the particles (WBC, RBC, PLT) to be examined, and at least one coating flow (MS) of separation step, which flows longitudinally of the central flow (ZS) through the flow passage (F1) of the separation stage (S1) and exhibits liquid (BL) to be examined, - an additional wave generation device (WE1) for piezoacoustic generation of sound waves (AW), which propagate through the passage (F1) of the separation stage (S1) transversely to the flow direction of the liquid (BL) to be examined flow and form a node of waves (KN) in a plane through the flow (ZS) central so that due to the pressure effect of sound waves (AW) in transverse direction , displaces particles (WBC, PLT) from the liquid (BL) to be examined to the flow (ZS) and they are concentrated there. Device according to claim 5, wherein the at least one separation passage lining flow (MS) surrounds the central flow (ZS) in a direction transverse to the flow direction and/or the liquid (PF ) initially free of the particles (WBC, RBC, PLT) to be examined, is transparent. Device according to claim 5 or 6, wherein the wave generating device (WE1, WE2) is designed such that the propagation direction of the sound waves (AW) in the separation stage (S1) and the propagation direction of the sound waves (AW) in the process stage (S2) run mutually orthogonal and orthogonal to the flow direction of the liquid (BL) to be examined. Device according to one of Claims 2 to 7, wherein the flow passage (F3) of the process step (S2) extends its width transversely to the flow direction, before the longitudinal position of the device (WE2 ) for generating waves of the process step (S2), so that the flow velocity of the liquid (BL) to be examined and of the at least one coating liquid (PF1, PF2) are reduced. Device according to one of Claims 1 to 8, having a buffer addition unit for adding buffer to the hypotonic solution in the central flow (ZS) or the fluid (BL) to be examined . Device according to one of Claims 1 to 9, which has a unit, which is preferably connected to the second input (E4), for the addition of markers for the particles (WBC, RBC, PLT) to be examined, such that the second coating liquid (PF2) comprises the markers. 11. Device for microscopic observation of particles (RBC, WBC) in a liquid to be examined, comprising: - a device (1, 2) according to one of claims 1 to 10, - a microscope (M), which is arranged in a longitudinal position of the flow passage (F) of the process step (S2), for the observation of particles (WBC, RBC, PLT) of the liquid to be examined in the plane (BA) of observation. Device according to claim 11, further exhibiting: - a collection unit (S3), which is connected in series with the processing stage (S2), downstream of the longitudinal position of the microscope (M) and is configured to collect the particles (WBC, RBC, PLT) examined, to examine these further with additional examination method, which requires an enrichment of the examined particles (WBC, RBC, PLT), preferably a molecular biology measurement method, and/or - a particle transfer unit, which is arranged downstream behind the longitudinal position of the microscope (M) and is configured for the transfer of particles (WBC, RBC, PLT) from the flow path (F) to a slide (OT ) for further examination of particles (WBC, RBC, PLT). 13. Procedure for the examination of particles (RBC, WBC, PLT) in a liquid (BL) to be examined, exhibiting the steps: - movement of a liquid (BL) to be examined with a first flow rate (Vs) through a flow path (F), - supply of at least one coating liquid (PF1, PF2) to the flow passage (F), through a first inlet (E3) with a second flow rate (V ³ ), so that the at least a coating liquid (PF1, PF2) forms at least one coating stream (MS1, MS2) in the flow passage (F) and the liquid (BL) to be examined flows through the coating passage (F). flow longitudinally with respect to the at least one coating flow (MS1, MS2), - piezoacoustic generation of sound waves (AW), which propagate through the flow passage (F) transversely to the flow direction of the liquid (BL) to be examined and formation of wave nodes (KN) in an observation plane (BA) so that, due to the pressure effect of the sound waves (AW) in the transverse direction, the particles (WBC, RBC, PLT) to be examined from the liquid (BL) to be being examined move transversally in the observation plane (BA) and there they are concentrated, - choice of the value of the at least one second flow rate (V ³ ), flow rate (V ⁴ ), so that the at least one coating flow (MS1, MS2) exhibits a predetermined cross section, of so that the observation plane (BA) runs through the at least one coating flow (MS1, MS2) and the particles (WBC, RBC, PLT) to be examined are concentrated in the at least one liquid ( PF1, PF2) lining, where by means of the arrangement of reflection elements opposite the wave generation device (WE2), the formation of standing waves is achieved, wherein drive elements generate the individual flows of the at least one coating liquid (PF1, PF2) and the liquid (BL) to be examined, wherein a control unit for the choice or control of the flow rates mutually predicts and synchronizes the values for the individual flow rates and controls the actuating element, to generate the individual flows of the coating liquids and of the liquid to be to be examined, corresponding to the determined flow rates. Use of a device (1, 2) according to one of claims 1 to 12 for the separation of leukocytes (WBC) from a blood plasma (BL).